It is widely believed that high density lipoprotein (HDL) protects against atherosclerosis by removing excess cholesterol from arterial cells. Recent studies indicate that HDL is also anti-inflammatory and inhibits lipid oxidation in vivo. These properties may contribute significantly to HDL's ability to inhibit atherosclerosis. Inflammation has been proposed to convert HDL to a dysfunctional form that loses these antiatherogenic effects. Understanding the role of dysfunctional HDL may lead to new diagnostic and therapeutic approaches to atherosclerosis and other inflammatory conditions. However, the underlying factors that render HDL dysfunctional remain poorly understood. One important pathway may involve oxidative damage to HDL by myeloperoxidase (MPO). We have shown that oxidation of apoA-l by MPO impairs the apolipoprotein's ability to remove cellular cholesterol by the ABCA1 pathway, and that HDL is targeted for damage by MPO in vivo. Furthermore, remarkably little is known regarding the identity of proteins that are carried in normal and dysfunctional HDL. We have used shotgun proteomics to test the hypothesis that HDL might carry proteins that make a previously unsuspected contribution to its cardioprotective activity. Our observations suggest that HDL carries a unique cargo of proteins in CVD subjects and that these proteins might make previously unsuspected contributions to the pro- and anti-inflammatory properties of HDL. We therefore propose to test the hypothesis that site-specific oxidation of apoA-l by MPO and deleterious alterations in the protein composition of HDL are molecular mechanisms for generating dysfunctional HDL in humans. Our specific aims are: i) Establish whether apolipoprotein A-l of HDL is targeted for oxidation in humans at risk for cardiovascular disease, ii) Determine whether site-specific oxidation of apoA-l in humans associates with loss of the apolipoprotein's ability to remove cellular cholesterol by the ABCA1 and ABCG1 pathways, iii) Determine whether pro- and anti-inflammatory proteins help generate dysfunctional HDL in humans.